Longart Marines, Liu Yun, Karavanova Irina, Buonanno Andres
Section on Molecular Neurobiology, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892, USA.
J Comp Neurol. 2004 Apr 26;472(2):156-72. doi: 10.1002/cne.20016.
Neuregulin-1 (NRG-1) regulates numerous aspects of neural development and synaptic plasticity; the functions of NRG-2 and NRG-3 are presently unknown. As a first step toward understanding how NRGs contribute to distinct aspects of neural development and function, we characterized their regional and subcellular expression patterns in developing brain. The expression of NRG-1-3 mRNAs was compared postnatally (P0, P7, adult) by using in situ hybridization. NRG-1 expression is highest at birth, whereas NRG-2 mRNA levels increase with development; expression of both genes is restricted to distinct brain regions. In contrast, NRG-3 transcripts are abundant in most brain regions throughout development. NRG-2 antibodies were generated to analyze protein processing, expression, and subcellular distribution. As with NRG-1, the transmembrane NRG-2 proprotein is proteolytically processed in transfected HEK 293 cells and in neural tissues, and its ectodomain is exposed and accumulates on the neuron surface. Despite the structural similarities between NRG-1 and NRG-2, we unexpectedly found that NRG-2 colocalizes with MAP2 in proximal primary dendrites of hippocampal neurons in culture and in vivo, although it is not detectable in axons or in axon terminals. These findings were confirmed with NRG-2 ectodomain antisera and epitope-tagged recombinant protein. In cerebellum, NRG-2 colocalizes with calbindin in proximal dendrites and soma of Purkinje cells. In contrast, NRG-1 is highly expressed in axons of dissociated hippocampal neurons, as well as in somas and dendrites. The distinct temporal, regional, and subcellular expression of NRG-2 suggests its unique and nonredundant role in neural function.
神经调节蛋白-1（NRG-1）调节神经发育和突触可塑性的多个方面；目前尚不清楚NRG-2和NRG-3的功能。作为理解NRGs如何促进神经发育和功能不同方面的第一步，我们对其在发育中的大脑中的区域和亚细胞表达模式进行了表征。通过原位杂交比较了出生后（P0、P7、成年）NRG-1-3 mRNA的表达。NRG-1的表达在出生时最高，而NRG-2 mRNA水平随发育增加；这两个基因的表达都局限于不同的脑区。相比之下，NRG-3转录本在整个发育过程中在大多数脑区都很丰富。生成了NRG-2抗体以分析蛋白质加工、表达和亚细胞分布。与NRG-1一样，跨膜NRG-2前体蛋白在转染的HEK 293细胞和神经组织中进行蛋白水解加工，其胞外结构域暴露并积聚在神经元表面。尽管NRG-1和NRG-2在结构上有相似之处，但我们意外地发现，NRG-2在培养和体内的海马神经元近端初级树突中与MAP2共定位，尽管在轴突或轴突终末中检测不到。这些发现用NRG-2胞外结构域抗血清和表位标记的重组蛋白得到了证实。在小脑中，NRG-2在浦肯野细胞的近端树突和胞体中与钙结合蛋白共定位。相比之下，NRG-1在解离的海马神经元的轴突以及胞体和树突中高表达。NRG-2独特的时间、区域和亚细胞表达表明其在神经功能中具有独特且不可替代的作用。
